A large number of monogenic human diseases are associated with genetic polymorphic variations or mutations in the so-called susceptibility genes. See generally, Cooper et al. in The Metabolic and Molecular Bases of Inherited Diseases, 1:259-291 (1995), Scriver et al., eds., McGraw-Hill, New York. One of the best-known and practically significant disease genes is the breast cancer susceptibility gene 1 (BRCA1), a tumor suppressor gene identified based on its genetic linkage to familial breast cancers. Mutations of the BRCA1 gene in humans are associated with predisposition to breast and ovarian cancers. In fact, BRCA1 and BRCA2 mutations are responsible for the majority of familial breast cancers. Inherited mutations in the BRCA1 and BRCA2 genes are responsible for approximately 7-10% of all breast and ovarian cancers. Women with BRCA mutations have a lifetime risk of breast cancer between 56% and 87%, and a lifetime risk of ovarian cancer between 27% and 44%.
With a large number of deleterious mutations identified in various disease susceptibility genes, genetic testing on patients to determine the presence or absence of such deleterious mutations proves to be an effective approach in detecting predispositions to diseases associated with such deleterious mutations. Indeed, genetic testing continues to grow in importance. For example, genetic testing is now commonly accepted as the most accurate method for diagnosing hereditary breast cancer and ovarian risk.
As is generally known in the art, humans are diploid, i.e., human autosomal genes are present in the genome in two copies. A mutation in one copy of a gene can be relevant even if the other copy of the gene is unaffected. This phenomenon is particularly notable in autosomal dominant genes such as BRCA1. However, most genetic testing approaches rely on the analysis of genetic materials amplified from patient samples that include a mixture of both gene alleles. The amplification techniques employed are typically indiscriminative of the two gene alleles in a diploid subject. For example, the most commonly utilized PCR-based genetic tests entail PCR amplification of different portions of both alleles of a gene and detecting mutations in those amplified portions by, e.g., sequencing or SNP detection, which may identify polymorphisms but cannot assign the identified variants to specific alleles. A more serious limitation inherent in such approaches is that they are not suitable for detecting genomic rearrangements (e.g., deletions or duplications) especially when a large rearrangement occurs in one but not the other allele. Because the techniques do not differentiate different alleles, if one allele is wild type and the other allele has a large deletion, the analysis result based on the techniques would show wild-type. The result misrepresents homozygosity as hemizygosity.
Mutations in many disease susceptibility genes are dominant mutations, i.e., mutations in only one allele of a patient are often sufficient to predispose the patient to diseases even if the other allele is wild type. This is especially true with large genomic rearrangements. Therefore, it is important to identify all mutations including large genomic: rearrangements. It will be particularly advantageous to complement traditional screening techniques that fail to distinguish between homozygous and hemizygous states with a method that can detect large genomic rearrangements.